Abstract Genetically modified mosquitoes resistant to Plasmodium development have been proposed as an alternative strategy to reduce malaria transmission. Several proof-of-principle studies have validated the idea that genetically modified mosquitoes can be refractory to the malaria parasite development, without incurring significant fitness loads. Recently, signaling cascades have been exploited to manipulate both parasite resistance and fitness. For example, manipulation of the insulin/insulin growth factor 1 signaling (IIS) pathway has been shown to confer Plasmodium resistance when up or down regulated via unique mechanisms, and extended lifespan when downregulated. However, the impact of IIS in other mosquito tissues has not been explored as extensively. The fat body of mosquitoes and other model invertebrates performs a number of functions, including storage of nutrients, production of yolk proteins and the synthesis of antimicrobial peptides. IIS has been implicated in these fat body processes leading to control of immunity, lifespan, metabolism, and reproduction. We previously generated a transgenic Anopheles stephensi line with increased insulin signaling in the peripheral fat body. Surprisingly, these transgenic mosquitoes survived significantly longer than their non-transgenic siblings, while in nearly every other organism and tissue increased IIS leads to a decrease in lifespan. To define how fat body IIS controls lifespan and to determine the impact fat body IIS has on reproduction, nutrient metabolism and Plasmodium resistance we will complete the following studies. Work in Drosophila suggest that fat body insulin signaling suppresses the expression of neuronal insulin-like peptides (ILPs) leading to increased lifespan. Thus, we will first examine transcript and peptide expression patterns of key AsILPs and knockdown putative AsILP targets via RNAi or Crispr knockout to validate the link between ILPs and lifespan extension. Our transgenic mosquito line also synthesized significantly more yolk protein than non-transgenic controls, although this did not translate into increased egg production during the first two gonotrophic cycles. Therefore, we will next conduct lifetime fecundity assays and development assays on the progeny to determine if an increase in lifetime reproductive fitness occurs. Third, due to the critical and well established role of IIS and the fat body on nutrient metabolism we will also quantify lipid, glycogen, glucose and trehalose levels at various physiological stages. Finally, the fat body is a key immune tissue regulating the production of anti-microbial peptides. As such we will assess the expression patterns of key immune genes and challenge transgenic mosquitoes with the most important human malaria parasite, Plasmodium falciparum. By the end of this project we will have a solid understanding of how fat body insulin signaling affects a range of physiologies impacting mosquito fitness and parasite resistance and will have developed new tools to generate highly fit Anopheles stephensi mosquitoes resistant to Plasmodium falciparum parasites.